The objective of the proposed research is to study the role of sterols in membrane and cellular functions using mutants of Neurospora which are defective in sterol biosynthesis. The phenotypes of the mutant studied so far differ from wild type with regard to growth, germination and sexual differentiation. These characteristics are apparently caused by the lack of wild type sterols (ergosterol and episterol) and/or the presence of abnormal sterols (precursors of ergosterol) in the cell membranes. Studies described in this proposal will be concerned with examining whether specific characteristics of the mutants are caused by unusual lipid-lipid interactions which affect membrane fluidity or if they result as a consequence of lipid-protein interactions. Initial experiments will involve characterization of lipids from functionally distinct membranes to determine the types and intracellular distribution of membrane lipid species. Effects of mutant sterols on membrane fluidity will be studied by observing the effects of rapid temperature shifts on membrane structure as monitored by freeze fracture electron microscopy. These studies will also include an investigation of the ability of the mutants to regulate membrane fluidity in acclimating to new growth temperatures. In order to determine the effects of mutant sterols on a specific membrane function, the amino acid transport capabilities of mutants with altered plasma membrane components will be examined. Following initial characterizations, experiments will be undertaken to distinguish between functions affected by sterol-lipid as opposed to sterol-protein interactions by determining how membrane lipid components affect various aspects of the mutant phenotype.